Abstract:Two sudanI hapten derivatives were synthesized by diazotization and the resulting products were characterized by liquid chromatography-mass spectrum. The haptens were conjugated with bovine serum albumin(BSA)and ovalbumin(OVA) respectively. UV spectrum and electrophoresis confirmed the successful conjugation. The BSA conjugates were used to immunize New Zealand rabbits for preparation of antibody while OVA conjugates were used as coating antigen. Indirect competitive ELISA methods were developed. Various factors including coating buffer,blocking solution,sample solution,antibody solution,reaction time and so on were optimized.Under the optimum conditions,an inhibition curve was established with IC50value of 3.0 ng/mL and linear range 0.3~23.4 ng/mL. The limit of detection(LOD) was calculated as 0.1 ng/mL. Crossreaction tests showed that the antibody recognized para red well with cross-reactivity of 140% while little recognition to sudan Ⅱ(1.1%),sudan Ⅲ(6.85%),sudan G(6.5%) and sudan Ⅳ(less than0.1%). Fortified chili powder of two levels(5 ng/g and 10 ng/g) were tested using the developed immunoassay. The recoveries were 90.4% and 96.0% with coefficients of variation 1.8% and 4.9%.
